The invention relates to the identification of expression profiles and the nucleic acids involved in colorectal cancer, and to the use of such expression profiles and nucleic acids in diagnosis and prognosis of colorectal cancer. The invention further relates to methods for identifying and using candidate agents and/or targets which modulate colorectal cancer.
Colorectal cancer is a significant cancer in Western populations. It develops as the result of a pathologic transformation of normal colon epithelium to an invasive cancer. There have been a number of recently characterized genetic alterations that have been implicated in colorectal cancer, including mutations in two classes of genes, tumor-suppressor genes and proto-oncogenes, with recent work suggesting that mutations in DNA repair genes may also be involved in tumorigenesis. For example, inactivating mutations of both alleles of the adenomatous polyposis coli (APC) gene, a tumor suppressor gene, appears to be one of the earliest events in colorectal cancer, and may even be the initiating event. Other genes implicated in colorectal cancer include the MCC gene, the p53 gene, the DCC (deleted in colorectal carcinoma) gene and other chromosome 18q genes, and genes in the TGF-xcex2 signalling pathway. For a review, see Molecular Biology of Colorectal Cancer, pp238-299, in Curr. Probl. Cancer, September/October 1997.
Imaging of colorectal cancer for diagnosis has been problematic and limited. In addition, dissemination of tumor cells (metastases) to locoregional lymph nodes is an important prognostic factor; five year survival rates drop from 80 percent in patients with no lymph node metastases to 45 to 50 percent in those patients who do have lymph node metastases. A recent report showed that micrometastases can be detected from lymph nodes using reverse transcriptase-PCR methods based on the presence of mRNA for carcinoembryonic antigen, which has previously been shown to be present in the vast majority of colorectal cancers but not in normal tissues. Liefers et al., New England J. of Med. 339(4):223 (1998).
Thus, methods that can be used for diagnosis and prognosis of colorectal cancer would be desirable. Accordingly, provided herein are methods that can be used in diagnosis and prognosis of colorectal cancer. Further provided are methods that can be used to screen candidate bioactive agents for the ability to modulate colorectal cancer. Additionally, provided herein are molecular targets for therapeutic intervention in colorectal and other cancers.
The present invention provides methods for screening for compositions which modulate colorectal cancer. In one aspect, a method of screening drug candidates comprises providing a cell that expresses an expression profile gene as set forth in FIG. 1 or a fragment thereof. The method further includes adding a drug candidate to the cell and determining the effect of the drug candidate on the expression of the expression profile gene.
In one embodiment, the method of screening drug candidates includes comparing the level of expression in the absence of the drug candidate to the level of expression in the presence of the drug candidate, wherein the concentration of the drug candidate can vary when present, and wherein the comparison can occur after addition or removal of the drug candidate. In a preferred embodiment, the cell expresses at least two expression profile genes. The profile genes may show an increase or decrease.
Also provided herein is a method of screening for a bioactive agent capable of binding to a colorectal cancer modulator protein (CCMP), the method comprising combining the CCMP and a candidate bioactive agent, and determining the binding of the candidate agent to the CCMP. Preferably the CCMP is a product encoded by a gene set forth in FIG. 1 SEQ ID NO:1 or a fragment thereof. Preferred fragments include any one or more of the peptides shown in FIG. 4. SEQ ID NOS:4-10.
Further provided herein is a method for screening for a bioactive agent capable of modulating the activity of a CCMP. In one embodiment, the method comprises combining the CCMP and a candidate bioactive agent, and determining the effect of the candidate agent on the bioactvity of the CCMP. Preferably the CCMP is a product encoded by a gene set forth in FIG. 1 SEQ ID NO:1 or a fragment thereof. In one embodiment, the CCMP is a peptide as shown in FIG. 3. SEQ ID NO:3 In another embodiment, the CCMP is a peptide as shown in FIG. 4. SEQ ID NO:4.
Also provided is a method of evaluating the effect of a candidate colorectal cancer drug comprising administering the drug to a transgenic animal expressing or over-expressing the CCMP, or an animal lacking the CCMP, for example as a result of a gene knockout.
Additionally, provided herein is a method of evaluating the effect of a candidate colorectal cancer drug comprising administering the drug to a patient and removing a cell sample from the patient. The expression profile of the cell is then determined. This method may further comprise comparing the expression profile to an expression profile of a healthy individual.
Moreover, provided herein is a biochip comprising a nucleic acid segment as set forth in FIG. 1 SEQ ID NO:1 or a fragment thereof, wherein the biochip comprises fewer than 1000 nucleic acid probes. Preferable at least two nucleic acid segments are included.
Furthermore, a method of diagnosing a disorder associated with colorectal cancer is provided. The method comprises determining the expression of a gene as set forth in FIG. 1 SEQ ID NO:1 or a fragment thereof in a first tissue type of a first individual, and comparing the distribution to the expression of the gene from a second normal tissue type from the first individual or a second unaffected individual. A difference in the expression indicates that the first individual has a disorder associated with colorectal cancer.
In another aspect, the present invention provides an antibody which specifically binds to CAA9, or a fragment thereof. In a preferred embodiment, the fragment of CAA9 is selected from the group consisting of CAA9p1, SEQ ID NO:4 CAA9p2, SEQ ID NO:4 CAA9p3, SEQ ID NO:4 CAA9p4, SEQ ID NO:5 CAA9p4MAPS, SEQ ID NO:6 CAA9p5 SEQ ID NO:7 and CAA9p5MAPS. SEQ ID NO:8. Preferably the antibody is a monoclonal antibody. The antibody can be a fragment of an antibody such as a single stranded antibody as further described herein, or can be conjugated to another molecule. In one embodiment, the antibody is a humanized antibody.
In one embodiment a method for screening for a bioactive agent capable of interfering with the binding of a colorectal cancer modulating protein (CCMP) or a fragment thereof and an antibody which binds to said CCMP or fragment thereof. In a preferred embodiment, the method comprises combining a CCMP or fragment thereof, a candidate bioactive agent and an antibody which binds to said CCMP or fragment thereof. The method further includes determining the binding of said CCMP or fragment thereof and said antibody. Wherein there is a change in binding, an agent is identified as an interfering agent. The interfering agent can be an agonist or an antagonist. Preferably, the antibody as well as the agent inhibits colorectal cancer.
In a further aspect, a method for inhibiting colorectal cancer is provided. In one embodiment, the method comprises administering to a cell a composition comprising an antibody to CAA9 or a fragment thereof. The method can be performed in vitro or in vivo, preferably in vivo to an individual. In a preferred embodiment the method of inhibiting colorectal cancer is provided to an individual with cancer. As described herein, methods of inhibiting colorectal cancer can be performed by administering an inhibitor of CAA9 activity. In one embodiment, a CAA9 inhibitor (used interchangeably with inhibitor of CAA9) is an antisense molecule to CAA9.
In another aspect of the present invention a method of eliciting an immune response in an individual is provided. In one embodiment, said method comprises administering to said individual a composition comprising CAA9 or a fragment thereof. In another embodiment said composition comprises a nucleic acid comprising a sequence encoding CAA9 or a fragment thereof.
Also provided herein is a composition capable of eliciting an immune response in an individual. In one embodiment, said composition comprising CAA9 or a fragment thereof and a pharmaceutically acceptable carrier. In another embodiment said composition comprises a nucleic acid comprising a sequence encoding CAA9 or a fragment thereof and a pharmaceutically acceptable carrier.
Other aspects of the invention will become apparent to the skilled artisan by the following description of the invention.